F/1F/O type ATPases are an important group of membrane-bound multisubunit enzymes involved in using ATP to generate a proton gradient, which can be used for substrate or ion transport. In mitochondria chloroplasts and in the plasma membrane of bacteria, the F/1F/O type ATPase also synthesizes ATP. We are studying the structure of the F/1F/O from Escherichia coli (ECF/1F/O) by a number of different approaches including cryoelectron microscopy, chemical cross-linking, protease digestion experiments, nuclear magnetic resonance studies, and fluorescence methods. We are also examining the conformational changes in ECF/1F/O that are involved in functioning of the enzyme complex. This aim is greatly aided by the fact that we can mutate subunits of the enzyme to include cysteine residues at defined positions in their sequences. These introduced cysteine residues are used to covalently attach reporter groups of conformational changes. In this way, kinetic analyses of conformational changes at various sites in the enzyme complex can be performed.